The permeability of red blood cells, with emphasis on the role of membrane electrostatics, will be studied. Recent research on artificial lipid bilayers shows the existence of huge electrostatic forces arising from surface potentials, dipole potentials, image forces, and the Born charging energy, that are normally present in any bilayer membrane. These forces are especially important in the partitioning and transport of small ions but must also play a significant role in the partitioning and conformation of proteins in natural membranes. Yet, there have been very few studies relating these forces to biological membrane function. These areas will be investigated by measuring and analysing the effects of systematic perturbations of transmembrane potentials, surface potentials, dipole potentials, and membrane dielectric constant. We will begin with transport through the lipid bilayer, where energy barriers will be used to interpret hydrophobic ion flux data (I-V) curves. Surface potentials will be monitored on both surfaces of resealed ghosts with EPR probes and with the permeation of lipid soluble ions. Dipole potentials will be estimated from the permeability ratios of hydrophobic anion and cation analogs. Membrane dielectric properties, altered by treatment with butanol and other reagents will be probed by measuring the permeability of hydrophobic ions of differing radii. Membrane composition will be altered by oxidation, cholesterol depletion/enrichment, and by altering symmetry. Similar techniques will be employed to attack problems of protein mediated transport. The surface potential on the inner surface will be characterized and its role in Ca sensitive membrane function as well as its role in producing asymmetric anion exchange characteristics will be assessed. Employing more than one probe, evidence for local heterogeneity of surface potential will be sought, and results used to characterize different ionic pathways. Effects of dipole potentials on urea and anion transport will be assessed by using different dipole reagents and correlating measured change in dipole potential with change in transport. A similar strategy will be employed to assess effects of changes in membrane dielectric constant. This is one of the first studies on effects of electrostatic lipid protein interactions on physiological transport. These interactions are disturbed during cell aging and pathology (e.g., exposure to oxidant stress such as ozone, drugs, and certain genetic diseases such as sickle cell anemia and NADPH deficiency).